Endothelins Regulations in ovarian Cancer

By: Mr.Raghavendra Basavaraja,By: Mr.Raghavendra Basavaraja, M.SC Animal sciences,M.SC Animal sciences, Faculta,Faculta,Hebrew university of Jerusalem Hebrew university of Jerusalem 04/10/23 Raghuippadi 1

Introduction Facts Papers Objectives Material & Methods Results Future strategies for OC

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Ovary Female reproductive organ Responsible for produce female sex

hormones and Ovulation

Cancer Abnormal growth of cells Caused by either Mutations or

abnormal activation of genes

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Some interesting facts about

OC

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Localized (15%)Confined to Primary SiteRegional (18%)Spread to Regional Lymph NodesDistant (61%)Cancer Has MetastasizedUnknown (6%)Upstaged

Percent of Cases by Stage

5-Year Relative Survival

Percent of Cases & 5-Year Relative Survival by Stage at Diagnosis: Ovary Cancer

Age Group: Ovary Cancer

Courtesy:SEER 18 2006-2010, All Races, Females

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EndothelinsEndothelins

A 21 amino acid peptide family ( EDN-1,2,3) Found in endothelial cells Pathophysiology….

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EDN’s PathwayEDN’s Pathway

ZibotentanZibotentan gefitinibgefitinib

Department of Animal Sciences, Faculty of Agricultural, Food and Environmental Quality Sciences, The Hebrew University of Jerusalem, Israel

Department of Animal Sciences, Faculty of Agricultural, Food and Environmental Quality Sciences, The Hebrew University of Jerusalem, Israel

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Expression of EDN’S In OC Epithelial cells. Cross talk b/w ETAR and EGFR enhances

the metasatic nature of OC Role of β-catenin & ECE-1 in metastasis & Role of β-catenin & ECE-1 in metastasis &

effect of their inhibitioneffect of their inhibition

Anticancer therapy Multi drug approach(Zibotetan+gefinib)

RNA interference

ETETAA-R-R & β-catenin & β-catenin

Endothelin-1Endothelin-1

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Materials and methods Materials and methods cell culture & Transfectioncell culture & Transfection: OVACR3(HTB61);OVACR3(HTB61);

ES2(CRL1978);EOC ;HEY & SKOV3ES2(CRL1978);EOC ;HEY & SKOV3

Real time PCRReal time PCR : CDNA-RNA( Gene expression)CDNA-RNA( Gene expression)

Western blot : Western blot : proteins Identification(MAPK;PKA)proteins Identification(MAPK;PKA)

ELISA & RIAELISA & RIA: ET-1 and β-catenin concentrationET-1 and β-catenin concentration

Immuno histochemistry: Immuno histochemistry: location of cellular proteinslocation of cellular proteins

Zymography: Zymography: protein activity (MMP)protein activity (MMP)

Statistical Analysis: Statistical Analysis: ± SEM & student t test( b/w groups)± SEM & student t test( b/w groups)

Multiple comparison by one way ANOVA & Tukey Kramer testMultiple comparison by one way ANOVA & Tukey Kramer test

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Results!!!

Results!!!

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Fig. 1. A. Lysates of SKOV3 cells treated with 10 ng/ml EGF, or 100 nM ET-1, and/or 1 μM zibotentan, or 1 μM gefitinib, or 1 μM PP2, or combination of zibotentan and gefitinib, were analysed by I...

Roberta Cianfrocca , Piera Tocci , Francesca Spinella , Valeriana Di Castro , Anna Bagnato , Laura Rosanò

The endothelin A receptor and epidermal growth factor receptor signaling The endothelin A receptor and epidermal growth factor receptor signaling converge on β-catenin to promote ovarian cancer metastasisconverge on β-catenin to promote ovarian cancer metastasis

Life Sciences Volume 91, Issues 13–14 2012 550 - 556

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Fig. 3. A. HEY cells were injected i.p. into nude mice and one week after cell injection, mice were treated for 21 days with vehicle, or with zibotentan (10 mg/kg...

Roberta Cianfrocca , Piera Tocci , Francesca Spinella , Valeriana Di Castro , Anna Bagnato , Laura Rosanò

Life Sciences Volume 91, Issues 13–14 2012 550 - 556

Effect of zibotentan and gefitinib treatments on metastatic potential of Effect of zibotentan and gefitinib treatments on metastatic potential of ovarian cancer cellsovarian cancer cells

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Levels of ET-1 mRNA (A), peptide concentrations in culture medium (B), and ECE-1 (C) and ETA Levels of ET-1 mRNA (A), peptide concentrations in culture medium (B), and ECE-1 (C) and ETA (D) mRNA expression in the ES2 and OVCAR3 ovarian carcinoma cell lines. (D) mRNA expression in the ES2 and OVCAR3 ovarian carcinoma cell lines.

Rayhman O et al. Cancer Res 2008;68:9265-9273

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Suppression of ET-1 biosynthesis by ECE-1 siRNA Suppression of ET-1 biosynthesis by ECE-1 siRNA molecules. molecules.

Rayhman O et al. Cancer Res 2008;68:9265-9273

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Effect of ECE-1 silencing on p44/42 MAPK phosphorylation.

Rayhman O et al. Cancer Res 2008;68:9265-9273

©2008 by American Association for Cancer Research

ConclusionConclusion ET-1 is one of major EDN’s express in the ovarian

cancer epithelial cells. ET-1 exerts its effects via transactivation of EGFR by

binding with ETAR. From the above pathway also we know that EDN can

activate β catenin to induce the metastasis Multi drug like zibotentan & geftinib is better rather

than using single approach ECE-1 Silencing is a prominent tool in control of

Endothelin-1expression in OC Finally we realized that even through EDN’s are

21 aa peptide play major role in OC.04/10/23 Raghuippadi 18

Future Strategies in Ovarian CancerFuture Strategies in Ovarian Cancer

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References

http://www.endothelins.com/Conferences/ http://seer.cancer.gov/statfacts/html/ovary.html. http://www.medicinenet.com/ovarian_cancer/article.htm .

http://www.ncbi.nlm.nih.gov/pubmed/20625315 Cianfrocca R, Tocci P, Spinella F, Di Castro V, Bagnato A, Rosanò L.Life

Sci. 2012 Oct 15;91(13-14):550-6. doi: 10.1016/j.lfs.2012.03.023. Epub 2012 Mar 28.PMID: 22480520 [PubMed - indexed for MEDLINE]

Rayhman O, Klipper E, Muller L, Davidson B, Reich R, Meidan R. Small interfering RNA molecules targeting endothelin-converting enzyme-1 inhibit endothelin-1 synthesis and the invasive phenotype of ovarian carcinoma cells. Cancer Res. 2008;68:9265–9273. [PubMed].

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